Phosphatase activity levels in pasteurized goats' milk

A study of the effect of pasteurization on goats' milk took place in order to compare the effectiveness of two pasteurization processes (low and high pasteurization) by assessing the procedure microbiologically and biochemically. The results revealed that both pasteurization processes were effective, showing microbial destruction of approximately 100%. With regard to the determination of phosphatase activity to monitor pasteurization, data in both cases were higher than the limit established for a well pasteurized milk, according to the method used. Machine milked milk showing a lower microbial content nearly achieved the standard level by batch pasteurization, 0.75 μg phenol/ml, but 2.5 μg phenol/ml by the high temperature short time (HTST) method. Phosphatase activity levels of 1.8 and 4.3 μg phenol/ml were the mean values obtained for low and high pasteurization respectively. Our results lead to the conclusion that the batch method for pasteurization of goats' milk is more efficient in terms of total bacterial count than the HTST method. Furthermore, the technique used for the determination of phosphatase activity does not seem to be adequate for pasteurization control of goats' milk, since in very rare situations were we able to attain the limit established.     

Optimization of sustained-release diltiazem formulations in man by use of an in-vitro/in-vivo correlation

In-vitro/in-vivo correlations (IVIVC) are useful for predicting in-vivo results from in-vitro data. An IVIVC has been used to optimize a hydrocolloidal-based matrix tablet designed to be bioequivalent to an existing once-daily diltiazem HC1 product (Dilacor XR 240mg; Rhone-Poulenc Rorer). Data from a preliminary formulation dosed to fasted and fed subjects were used to establish the IVIVC. The correlation was then used during reformulation of the dosage forms to predict changes in the maximum plasma concentration (Cmax) and the area under the plasma-concentration-time curve (AUC) for fasted and fed subjects using in-vitro dissolution data. The IVIVC adequately predicted plasma profiles of two optimized formulations in studies with fasted and fed subjects.     

Interactions between imidazoline compounds and sulphonylureas in the regulation of insulin secretion

1. Imidazoline alpha 2-antagonist drugs such as efaroxan have been shown to increase the insulin secretory response to sulphonylureas from rat pancreatic B-cells. We have investigated whether this reflects binding to an islet imidazoline receptor or whether alpha 2-adrenoceptor antagonism is involved. 2. Administration of (+/-)-efaroxan or glibenclamide to Wistar rats was associated with a transient increase in plasma insulin. When both drugs were administered together, the resultant increase in insulin levels was much greater than that obtained with either drug alone. 3. Use of the resolved enantiomers of efaroxan revealed that the ability of the compound to enhance the insulin secretory response to glibenclamide resided only in the alpha 2-selective-(+)-enantiomer; the imidazoline receptor-selective-(-)-enantiomer was ineffective. 4. In vitro, (+)-efaroxan increased the insulin secretory response to glibenclamide in rat freshly isolated and cultured islets of Langerhans, whereas (-)-efaroxan was inactive. By contrast, (+)-efaroxan did not potentiate glucose-induced insulin secretion but (-)-efaroxan induced a marked increase in insulin secretion from islets incubated in the presence of 6 mM glucose. 5. Incubation of rat islets under conditions designed to minimize the extent of alpha 2-adrenoceptor signalling (by receptor blockade with phenoxybenzamine; receptor down-regulation or treatment with pertussis toxin) abolished the capacity of (+)- and (+/-)-efaroxan to enhance the insulin secretory response to glibenclamide. However, these manoeuvres did not alter the ability of (+/-)-efaroxan to potentiate glucose-induced insulin secretion. 6. The results indicate that the enantiomers of efaroxan exert differential effects on insulin secretion which may result from binding to effector sites having opposite stereoselectivity. Binding of (-)-efaroxan (presumably to imidazoline receptors) results in potentiation of glucose-induced insulin secretion, whereas interaction of (+)-efaroxan with a second site leads to selective enhancement of sulphonylurea-induced insulin release.     

The entry of [D-penicillamine2,5]enkephalin into the central nervous system: saturation kinetics and specificity

The delta opioid receptor-selective, enzymatically stable peptide [D-Penicillamine2,5]enkephalin (DPDPE) has recently acquired special significance with the identification of a saturable uptake system for this analgesic into the CNS. The aim of the present study was to characterize further the entry of [3H]DPDPE into the brain and CSF by means of a bilateral in situ brain perfusion method. Initial experiments revealed a saturable [3H]DPDPE uptake into the brain that followed Michaelis-Menten type kinetics with a K(m) value of 45.5 +/- 27.6 microM, a V(max) value of 51.1 +/- 13.2 pmol x min(-1) x g(-1) and a K(d) value of 0.6 +/- 0.3 microl x min(-1) x g(-1). Uptake of [3H]DPDPE into the CSF could not be inhibited (K(d) = 0.9 +/- 0.1 microl x min(-1) x g(-1)). Entry of [3H]DPDPE into the CNS was not inhibited in the presence of 10 mM 2-aminobicyclo-[2,2,1]-heptane-2-carboxylic acid (BCH) or 50 microM ICI 174,864, which suggests that the saturable mechanism does not involve the large neutral amino acid transporter or binding to opioid receptors. It would also appear that [3H]DPDPE is not in competition with either poly-L-lysine or insulin to enter the CNS. However, both of these substances significantly increased the CNS entry of [3H]DPDPE but not that of the vascular space marker [14C]sucrose, and this may have valuable clinical implications. It is not known at present which saturable uptake mechanism is responsible for the CNS entry of [3H]DPDPE, but overall the results suggest a carrier-mediated transport system.     

Replication initiation point mapping

Boric acid (H3BO3) has been used in a wide variety of applications--medication, pesticides, and household products. Reports of child poisoning by H3BO3 were common in the clinical literature before 1975. However, a decline in its use as a bacteriostatic agent coupled with increased regulatory control has almost eliminated poisonings by accidental ingestion. Schedule I (Part I, Item 8) of the Hazardous Products Act of Canada, proclaimed in the late 1960s, followed in the wake of concerns about accidental poisoning and prohibits its use in toys. Since that time, scientific knowledge has increased and has led to a reevaluation of the hazard associated with H3BO3. A maximum tolerated dose (MTD) was sought for children in the most susceptible age range, with a view to determine a maximum acceptable concentration (MAC) in toys. The effects of H3BO3 in a variety of exposure scenarios were evaluated. Precedence was given to clinical data in humans, particularly children, since there is no suitable animal model of boric acid intoxication. An extensive search of the pediatric literature was conducted to find a no-observed-adverse-effect level (NOAEL) or a lowest-observed-adverse-effect level (LOAEL). An analysis of the pivotal study to the present assessment resulted in the application of an uncertainty factor of 100 to account for variations in sensitivity among children and for the use of a LOAEL. Based on a pediatric LOAEL of 300 mg/kg body wt, we derived a MTD of 3 mg H3BO3/ kg body wt and a MAC of 9.1 mg H3BO3/g of toy. These results compared favorably with calculations from other human and animal NOAELs/LOAELs.     

Highly attenuated modified vaccinia virus Ankara (MVA) as an effective recombinant vector: a murine tumor model

Until recently, the majority of HLA class I typing has been performed by serology. Expensive commercial typing trays are frequently used for testing non-Caucasian subjects and new strategies using DNA-based methods have been adopted for improving clinical histocompatibility testing results and adapted as supplements in proficiency testing. A double-blind comparison of the typing of HLA-B specificities in 40 samples was carried out between serology and two polymerase chain reaction (PCR) methods, PCR amplification with sequence-specific primers (PCR-SSP) and PCR amplification and subsequent hybridization with sequence-specific oligonucleotide probes (PCR-SSOP). The results demonstrated 22.5% misassignments of HLA-B antigens by serology. There was complete concordance between the results obtained with the two PCR based typing methods. A second panel of 20 donor samples with incomplete or ambiguous serologic results was analyzed by PCR-SSP and SSOP Both PCR methods identified correctly the HLA-B antigens. Our results suggest that more accurate typing results can be achieved by complementing serologic testing with DNA-based typing techniques. The level of resolution for HLA-B antigen assignment can be obtained by this combination of serology and limited DNA-based typing is equivalent to the HLA-B specificities defined by the WHO-HLA Committee. This level of resolution cannot routinely be achieved in clinical histocompatibility testing or in proficiency testing using serologic reagents only.